Strong binding of drugs and biologically active substances to their target biopolymers is required for the appearances of their bioactivities. The biopolymers include pharmacological receptors which take part in signal transmission between cells, as well as enzymes, cytokines and other proteins, complexes comprising these proteins as main components, and nucleic acids. Since 1960, the steric structures of a large number of biopolymers have been revealed at the atomic level by X-ray crystallographic analyses and the structural data thereof have been stored in the Protein Data Bank and published.
Small molecules which can be bound to these biopolymers are called ligand molecules. The ligand molecules include candidate molecules for drugs such as drug molecules to be synthesized in future, as well as drugs, substrates and inhibitors of enzymes, coenzymes and the like.
For the stable binding between the biopolymers and ligand molecules it is required that the shapes of cavities which exist in the steric structures of biopolymers and into which ligand molecules are bound (hereinafter referred to as "ligand-binding sites") should be complementary to the shapes of the ligand molecule surfaces as in the case of locks and keys and that there should be interactions between the both molecules which result in specific affinity therebetween. It is known from the crystallographic analyses of biopolymer-ligand molecule complexes that among these intermolecular interactions, hydrogen-bond, electrostatic and hydrophobic interactions and the like are particularly important. In drug design and structure-activity relationship study, it is very important to know whether a compound molecule can form a stable complex with its target biopolymer and, if so, what kind of binding mode the complex has (i.e., which functional group in the ligand-binding site of the biopolymer interact which functional groups in the ligand molecule in what kind of mode) and how stable the complex is. In conformationally flexible ligand molecules, it is also important for drug design to determine the conformation of the ligand molecules bound to target biopolymers (active conformation) and such a determination must be made at the same time as searching stable binding modes.
It is known that when the complexes have the most stable structures in terms of free energy, the ligand molecule structures are often different from the crystal structures of unbound ligand molecules, the structures in solutions, and the energically most stable (or any local minimum) structures obtained by various kinds of energy calculations.
It is impossible to determine the modes of binding of all ligand molecules of interest to their target biopolymer by experimental methods such as X-ray crystallographic analysis. First, this is because a great effort and time are necessary for analyzing the mode of binding of each ligand molecule to the biopolymer by experimental methods. In addition, if the ligand molecules are substrates for enzymes, any stable structures of the complex can not be detected in the experiments due to the progress of enzymatic reactions. Furthermore, the samples of the ligand compounds are often unavailable if they exist. In some cases, it may be required to predict the binding mode and the stability of the complex between an imaginary ligand molecule and its biopolymer, in order to decide whether it is valuable to synthesize the ligand molecule. Such a prediction is very useful in making new drugs and biologically active substances by molecular design.
The simulation study of the mode of stable binding of ligand molecules to their target biopolymer whose structure is known is called "docking study". In the past, the docking study was performed using molecular models, but the use of molecular models had problems in that; it needed much time and effort to construct the molecular models, that the precision of the molecular models and the reproducibility of results were poor and that quantitative results could not be obtained. As a method for solving these problems, a simulation method using a computer and computer graphics is generally used today.
In the simulation method using a computer and computer graphics, it is the most common procedure that initial structures of complexes are roughly set interactively on computer graphics displays by visual decision and then they are refined and quantitated by computation methods. However, there are an enormous number of possible docking structures, and therefore it is very difficult to reach objectively the correct solution. Furthermore, it is a time-consuming and laborious method. In addition, the obtained results are unreliable and unreproducible in that they vary with users.
In order to solve these problems, a few methods in which user's preconception can be eliminated have been studied. Kuntz et al. published a program for approximately representing ligand molecules by sets of spheres and changing the relative positions between the spheres instead of changing the molecular conformations to determine whether the ligand molecules fit the ligand-binding site in a receptor (Docking Flexible Ligands to Macromolecular Receptors by Molecular Shape, R. L. Desjarlais, R. P. Sheridan, J. Scott Dixon, I. D. Kuntz and R. Venkataraghavan: J. Med Chem. (1986) 29, pp.2149-2153). However, this program is not effective in determining the binding modes and the ligand conformations in stable complexes in a reliable manner because it does not deal with specific intermolecular interactions such as hydrogen-bond and the like.
F. Jiang et al. developed an automatic docking method in which intermolecular interactions such as hydrogen-bond, electrostatic interaction and the like are considered qualitatively in addition to the shape complementarity which Kuntz et al. highlighted (F. Jiang and S. Kim, J. Mol. Biol. 219, 79 (1991)). However, this method neither considers the conformational flexibility of ligand molecules nor makes a sufficient quantitative estimate of intermolecular interactions.
None of other methods are effective because they have problems in that they do not consider specific intermolecular interactions or the conformational flexibility of ligand molecules. The binding mode of ligand molecules to biopolymers and the ligand conformations are coupled completely. Since the binding mode and active conformation have enormous numbers of possibilities, all the combinations thereof must be examined in order to obtain the most stable structure of the complex covering all possible binding modes and ligand conformations. However, no methods for searching stable structures of complexes that take into account all the combinations of the binding modes and ligand conformations have been established.
Therefore, an object of the present invention is to provide methods for searching stable docking models of biopolymer-ligand molecule complex that solve the above-mentioned problems.
Another object of the present invention is to provide methods for searching the modes of binding of ligand molecules to biopolymers and the active conformations of the ligand molecules at the same time.